Embedded Human Periodontal Ligament Stem Cells Spheroids Enhance Cementogenic Differentiation via Plasminogen Activator Inhibitor 1

• Published in: International journal of molecular sciences Vol. 23; no. 4; p. 2340
• Main Authors: Yasunaga, Madoka, Ishikawa, Hiroyuki, Tamaoki, Sachio, Maeda, Hidefumi, Ohno, Jun
• Format: Journal Article
• Language: English
• Published: Switzerland MDPI AG 20-02-2022; MDPI
• Subjects: Adipocytes; Biomedical materials; Cell culture; Cell Differentiation; Cells, Cultured; Cementogenesis; cementogenic differentiation; Cementum; Cementum attachment protein; cementum protein 1; Collagen; collagen embedding culture; Differentiation; Extracellular matrix; human periodontal ligament stem cells; Humans; Immunocytochemistry; Inhibitors; Ligaments; Monolayers; Morphology; Periodontal Ligament; Periodontium; plasminogen activator inhibitor 1; Plasminogen Activator Inhibitor 1 - genetics; Plasminogen Activator Inhibitor 1 - metabolism; Plasminogen activator inhibitors; Proteins; Proteins - metabolism; Spheroids; Spheroids, Cellular - metabolism; Stem cells; Stem Cells - metabolism; Tissue engineering; Tooth root; collagen embedding culture; human periodontal ligament stem cells; cementogenic differentiation; spheroids; cementum protein 1; plasminogen activator inhibitor 1
• ISSN: 1422-0067; 1661-6596; 1422-0067
• DOI: 10.3390/ijms23042340

Spheroids reproduce the tissue structure that is found in vivo more accurately than classic two-dimensional (2D) monolayer cultures. We cultured human periodontal ligament stem cells (HPLSCs) as spheroids that were embedded in collagen gel to examine whether their cementogenic differentiation could be enhanced by treatment with recombinant human plasminogen activator inhibitor-1 (rhPAI-1). The upregulated expression of cementum protein 1 (CEMP1) and cementum attachment protein (CAP), established cementoblast markers, was observed in the 2D monolayer HPLSCs that were treated with rhPAI-1 for 3 weeks compared with that in the control and osteogenic-induction medium groups. In the embedded HPLSC spheroids, rhPAI-1 treatment induced interplay between the spheroids and collagenous extracellular matrix (ECM), indicating that disaggregated HPLSCs migrated and spread into the surrounding ECM 72 h after three-dimensional (3D) culture. Western blot and immunocytochemistry analyses showed that the CEMP1 expression levels were significantly upregulated in the rhPAI-1-treated embedded HPLSC spheroids compared with all the 2D monolayer HPLSCs groups and the 3D spheroid groups. Therefore, 3D collagen-embedded spheroid culture in combination with rhPAI-1 treatment may be useful for facilitating cementogenic differentiation of HPLSCs.